Bryozoan Growth and Environmental Reconstruction by Zooid Size Variation

Vol. 430: 133–146, 2011 MARINE ECOLOGY PROGRESS SERIES Published May 26 doi: 10.3354/meps08965 Mar Ecol Prog Ser

Contribution to the Theme Section ‘Evolution and ecology of marine biodiversity’ OPENPEN ACCESSCCESS Bryozoan growth and environmental reconstruction by zooid size variation

Beth Okamura1,*, Aaron O’Dea2, 3, Tanya Knowles4

1Department of Zoology, Natural History Museum, London SW7 5BD, UK 2Center for Tropical Paleoecology and Archeology, Smithsonian Tropical Research Institute, Apartado 0843-03092 Panamá, República de Panamá 3Department of Earth and Ocean Sciences, University of British Columbia, Vancouver, British Columbia V6T 1Z4, Canada 4Department of Earth Science and Engineering, Imperial College London, London SW7 2BP, UK

ABSTRACT: The modular growth of cheilostome bryozoans combined with temperature-induced variation in module (zooid) size has enabled the development of a unique proxy for deducing seasonal temperature regimes. The approach is based on measures of intracolonial variation in zooid size that can be used to infer the mean annual range of temperature (MART) experienced by a bryozoan colony as predicted by a model of this relationship that was developed primarily to infer palaeosea- sonal regimes. Using the model predictions effectively requires a highly strategic approach to characterise the relative amount of within-colony zooid size variation (by adopting random or very systematic measurements of zooids that meet a stringent set of criteria) to gain insights on temperature variation. The method provides an indication of absolute temperature range but not the actual temperatures experienced. Here we review the development of, support for and applications of the zooid size MART approach. In particular, we consider the general issue of why body size may vary with temperature, studies that validate the zooid size–temperature relationship and insights that have been gained by application of the zooid size MART approach. We emphasise the potential limitations of the approach, including the inﬂuence of confounding factors, and highlight its advantages relative to other proxies for palaeotemperature inferences. Of prime importance is that it is relatively inexpensive and quick and allows a direct estimate of temperature variation experienced by an individual colony. Our review demonstrates a strong and growing body of evidence that the application of the zooid size MART approach enables robust interpretations for palaeoclimates and merits broad recognition by environmental and evolutionary biologists and climate modellers.

KEY WORDS: Cheilostomes · Mean annual range of temperature · MART · Body size–temperature relationship · Palaeoclimates

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INTRODUCTION continuous record of growth and the sequential development of discrete and measurable features that vary Patterns of growth in plants and animals have long consistently with respect to a single environmental been used to gain insights into past environments. variable and remain ﬁxed, thereby permitting the Variable accretion of structural material in trees, ﬁsh retrieval of environmental conditions relevant to par- otoliths and the shells of bivalve or gastropod molluscs, ticular time periods. Benthic colonial invertebrates for example, can be used to retrospectively extract can provide an especially appropriate system for such ambient environmental conditions such as rainfall, retrieval, since, with some exceptions (e.g. sponges), temperature or food availability (e.g. Falcon-Lang they comprise distinct, individual modules (zooids) that 2005, Zazzo et al. 2006, Hallmann et al. 2009). Organ- are produced iteratively throughout the lifetime of the ismal attributes that favour such analyses include a colony. The sclerochronological analysis of modular

*Email: [email protected] © Inter-Research 2011 · www.int-res.com 134 Mar Ecol Prog Ser 430: 133–146, 2011

growth can provide inferences for both intra- and 1989). Once the skeletal walls of a new zooid are interannual environmental variation, information that secreted, there is no further expansion of zooid surface is not readily available from analyses of short-lived, area (O’Dea & Okamura 2000b). This gives the zooid a unitary organisms that are commonly used as proxies, determinate size that has been shown to be controlled such as foraminifers or ostracodes. Furthermore, colo- to a signiﬁcant extent by the ambient water tempera- niality is often associated with polymorphism, with ture at the time the zooid was produced. Bryozoan modules specialised for different functions within a colonies therefore record the range of temperatures colony. These attributes, viz. modular iteration, poly- experienced during their lifetime as intracolonial vari- morphism and individual colony longevities ranging ation in zooid size (Fig. 1). Such temperature-induced from months to many years, may enable joint insights variation in size is also observed in unitary animals and into environmental conditions and associated life his- is generally known as the ‘temperature–size rule’ tory variation (O’Dea & Okamura 2000a, O’Dea & (Atkinson 1994). Jackson 2002). Such insights are generally difﬁcult to The above-mentioned features make cheilostome achieve through investigations of longer-lived unitary bryozoans unique amongst colonial taxa in offering organisms, such as bivalves or brachiopods, since the opportunities for inferring environmental conditions morphologies of these organisms do not readily pro- and biotic responses in the present day as well as over vide a record of functional allocation during their life- geological time. Other colonial taxa such as corals, time. Surprisingly, however, the unique contribution of hydroids, ascidians and other non-cheilostome bryo- colonial invertebrates for retrospectively deducing zoans either do not produce carbonate skeletons, show environmental and life history variation has not been indeterminate growth of their polyps or zooids, or ex- widely recognised. hibit little to no polymorphism and therefore preclude Bryozoans are colonial, suspension-feeding inverte- gaining additional insights on how life histories may brates that are common members of benthic assem- respond to environmental conditions. blages (McKinney & Jackson 1989). There are some Recognition of the unique opportunities afforded by 6000 described extant species of bryozoans (Gordon cheilostome bryozoans for the retrieval of environmen- et al. 2009), most of which belong to the order tal information led to the development of a method that Cheilostomata. Colonies of cheilostomes comprise allows the estimation of the mean annual range of asexually budded zooids that are reinforced by skele- temperature (MART) based on variation in zooid size tal walls composed of calcitic and/or aragonitic car - within cheilostome colonies. The method is based on bonate (Rucker & Carver 1969, Smith et al. 2004). using model predictions for how zooid size varies with Typically, cheilostomes display zooid polymorphism MART and thus requires that zooids meet a stringent (McKinney & Jackson 1989). The majority of zooids are set of criteria, in keeping with assumptions of the specialised for feeding (autozooids), whilst a smaller model, and that a strategic sampling protocol is adopted proportion function in reproduction (ovicells) and to target appropriate zooids randomly or very system- defense (avicularia). The carbonate skeleton (‘zooe- atically. The method informs on absolute seasonal vari- cium’) confers preservation of colony features, includ- ation in temperature but does not indicate the actual ing zooid polymorphism, and bryozoans are well rep- temperatures. Thus polar and tropical bryozoans will resented in the fossil record (McKinney & Jackson converge on similar low MART values.

Fig. 1. Cupuladria exfragminis. Sea- sonal variation in zooid size. Scanning electron micrographs of a recent col - ony from the Gulf of Panama. Size difference between zooids that developed during (A) upwelling (cold) and (B) non-upwelling (warm) conditions. Same magnification for the purpose of comparison. The skeletal walls of both autozooids (large orifices) and avicularia (small orifices) are evident. Scale bar = 150 µm. Photos by R. Dewel Okamura et al.: Environmental reconstruction by zooid size analysis 135

In this paper we describe this zooid size MART cent review). Mechanistic explanations have in cluded: approach, review studies that validate the zooid (1) the production of smaller adult stages because size–temperature relationship on which the technique developmental rate is more strongly influenced by in - depends and summarise research that has made use of creasing temperature than growth rate (van der Have the approach in order to demonstrate the insights that & de Jong 1996); (2) the related hypotheses that cell can be gained by its adoption. We also emphasise the (van Voorhies 1996, Woods 1999) or body (Chapelle & potential limitations of the zooid size MART approach, Peck 1999) size is limited by oxygen diffusion. describe confounding factors that must be borne in Atkinson et al. (2006) recently addressed the ex- mind and suggest directions for future studies. How- planation that the temperature–size relationship may ever, because the approach is based on temperature- relate to oxygen concentrations by examining the ther- induced variation in zooid size, we first describe the mal responses of the bryozoan Celleporella hyalina to temperature–size rule and address the mechanism(s) 2 temperatures (10 and 18°C) and 2 oxygen concentra- that may underlie the temperature–size relationship. tions (21 and 10%, representing normoxia and hypo- Examination of these issues leads us to conclude that xia, respectively). They found that smaller zooids were the zooid size MART approach provides a unique and produced under hypoxia regardless of temperature independent proxy that will enable more robust inter- (although size was also influenced directly by temper- pretations when incorporated as part of the toolkit ature), providing evidence for the expected adjustment used for environmental and evolutionary studies. of size in response to oxygen requirements. This adjustment is anticipated because increasing temperatures increase metabolic rates and thus oxygen de- VARIATION IN BODY SIZE WITH TEMPERATURE mands, but these metabolic oxygen demands increase faster with temperature than diffusion in the organ- The inverse relationship between zooid size and ism’s oxygen uptake and transport system. Smaller size temperature conforms to a general pattern observed will therefore decrease diffusion distances and in- in ectotherms known as the ‘temperature–size rule’ crease the relative surface to volume ratios for oxygen (Atkinson 1994). This pattern is expressed as pheno- uptake. In addition, size variation may regulate respi- typic plasticity in response to temperature variation ratory activity, with a decrease in size reducing activity demonstrated by negative thermal reaction norms. As at higher temperatures via reductions in mitochondrial with any ‘rule’, there are instructive exceptions, but volume density and in cristae density (see review by the overall weight of evidence for the temperature– Atkinson et al. 2006). size rule (Atkinson 1994, Angilletta et al. 2004, King- Atkinson et al. (2006) also obtained evidence that the solver & Huey 2008) lends additional and important temperature–size rule is not a fundamental response support for temperature-driven variation in zooid sizes. of cells. Larval parenchyma cells were larger at the What has been unclear, however, is what mechanisms lower temperature, but temperature had no effect on may underlie this nearly universal relationship, whether the size of epithelial cells of the tentacle. Similar there exists an adaptive basis for thermal sensitivity in results have been obtained in other empirical studies body size, and whether the mechanisms and/or adap- that have shown that larger body sizes at lower tem- tive explanations are inclusive across taxa. Central to peratures can be caused by an increase in cell size in this issue is the association of the temperature–size some systems (e.g. in nematodes or some Drosophila rule with a life-history puzzle (Berrigan & Charnov populations) but not in others (including other Droso - 1994): that good conditions result in faster growth to a phila populations; see Angilletta et al. 2004 for review). larger size but that temperature has contradictory As mentioned earlier, Atkinson et al. (2006) also found effects on growth and size, with higher temperatures that an inverse temperature–size relationship charac- driving faster growth to a smaller size. terised zooids, but this was not the case for tentacle Hypotheses proposed for the temperature–size rule length. Thus, the temperature–size rule did not apply include both adaptive and non-adaptive scenarios (At - universally at the cell or organ (e.g. tentacle) level. kinson 1994, Angilletta et al. 2004). However, demon- Their results led Atkinson et al. (2006) to propose strations that the shapes of thermal reaction norms can that temperature-induced size changes at different readily evolve in response to selection (see Kingsolver levels of organisation are part of a range of acclimation & Huey 2008 for review) suggest that selection main- mechanisms, including variation in body size, that will tains the temperature–size rule. A problem common to optimise functional capacity (e.g. of mitochondria and many of the hypotheses proposed to explain the tem- tissues) to maintain scope for aerobic activity. They perature–size rule is that they are too restrictive to suggest that these acclimatory processes occur within apply to the diversity of taxa that follow the rule (see a temperature range whose limits are determined by Angilletta et al. 2004, Kingsolver & Huey 2008 for re - when oxygen partial pressures of body fluids fall. This 136 Mar Ecol Prog Ser 430: 133–146, 2011

occurs at the so-called ‘pejus’ limits, when the capacity zooid size and the MART with 95% confidence limits of oxygen supply mechanisms is unable to support within ±1°C across the entire temperature range (see oxygen demand (Pörtner 2002). Thus, at high tempera- Fig. 2c in O’Dea & Okamura 2000b). Zooid size was tures, excessive oxygen demand results in insufficient estimated by measuring the maximum distance oxygen levels in the body fluids, while at low tempera- between the proximal and distal skeletal walls, to esti- tures, the aerobic capacity of mitochondria may be - mate zooid length, and the maximum distance be - come limiting. Beyond these limits aerobic scope dis- tween the lateral skeletal walls, to estimate zooid appears, and the adoption of anaerobic metabolism width (Fig. 2). These maximum distances infer growth will support time-limited survival. If these acclimatory along a straight line trajectory aligned perpendicular responses to maintain scope for growth underlie the to zooid margins that are farthest apart (see Fig. 2). temperature–size rule, they could at least in part ac- Zooid frontal area is then calculated as the product of count for the puzzle of the inverse temperature–size zooid length and width. The mean maximal difference relationship despite increased growth rate with warmer between summer and winter temperatures over a temperatures. number of years for the depths at which the bryozoans As discussed below and exemplified by the study of were collected (see O’Dea & Okamura 2000b for fur- Atkinson et al. (2006), cheilostome bryozoans dem- ther discussion) was used to estimate MART. Algebraic onstrate intraspecific thermal sensitivity that reflects rearrangement of the regression provides a means of the temperature–size rule. They also demonstrate vari- predicting MART as: MART = –3 + 0.745b, where b is ation in zooid size within (see ‘Evidence for relationship the mean intracolonial CV of zooid size. between temperature and zooid size’) and among The dependability of the zooid size MART approach closely related species (e.g. species of Haplopoma; is a function of several factors, including the data that Ryland 1963) living in geographic regions that are char- were originally used to develop the model, variation in acterised by different temperature regimes. The extent response to MART amongst taxa, other agents of zooid to which the latter reflects phenotypic plasticity versus size variation and rigour in applying the approach. selection requires investigations of thermal reaction In developing the model, data on zooid sizes were norms. It should be noted that the zooid size approach required to meet a stringent set of criteria. Zooid size to MART is not complicated by these issues since it is values were based on 20 randomly selected autozooids based on intracolonial zooid size variation. However, per colony. However, to minimise variation due to fac- the occurrence of the temperature–size rule at the tors known to influence zooid size, the autozooids were modular level amongst colonial organisms raises the required to show uninhibited growth (i.e. were not question of the adaptive significance of body size when perceptibly deformed), to be part of the basal series of environments change over the lifetime of colonies. Be- zooids (i.e. not frontally budded) and to be located out- cause zooids remain fixed in size, any adaptive basis for side of the zone of astogenetic change (Fig. 3). This size at the time of budding will be ephemeral in sea- zone is created during the early growth of colonies, sonal environments. For bryozoans, an adaptive basis when zooid generations show pronounced incremental for phenotypic plasticity in zooid size may nevertheless increases in size. Beyond this zone zooid sizes are rel- apply if at least some regions of a colony are in optimal atively stable (Boardman & Cheetham 1969). There condition as a result of zooid-size matching to the pre- were also criteria for choosing colonies and species for vailing thermal regime. Alternatively, smaller zooids measurement. For colonies, these included offering at may simply result if the developmental rate is more least 30 ontogenetically complete autozooids and strongly influenced by increasing temperature than avoiding measuring colonies whose shapes were com- growth rate (van der Have & de Jong 1996). promised by e.g. irregularities in the substratum or observable competition from other organisms. For species these included: offering clearly delimited zooid ZOOID SIZE MART APPROACH margins for measurement; no evidence of distortion in dried material; avoidance of ‘spot’ colonies that undergo The zooid size MART approach is based on a predic- determinate growth to a very small size (Winston & tive model that allows seasonal variation in tempera- Håkansson 1986, Bishop 1989); the availability of at ture regimes to be estimated from the empirically least 5 replicate colonies per species. The CVs were derived relationship between intracolonial zooid size averaged for each species at each locality. Since the variation and the MART (O’Dea & Okamura 2000b). model uses estimates of zooid size to infer seasonal The model was developed by undertaking morphome- temperature regimes, it is important that, as far as pos- tric analyses of 157 colonies of 29 cheilostome species sible, these estimates are based on applying a similar ranging from tropical to polar regions and is based on set of criteria to those that were used to develop the a regression of the mean coefficient of variation (CV) of model. Okamura et al.: Environmental reconstruction by zooid size analysis 137

somewhat smaller zooids that occur at bifurcations or just distal to bifurcations in the normal budding series. Such zooids are encountered more frequently near the centre of the colony, as row bifurcations allow zooids to populate space as the colony extends radially. In such cases, the path of the profile was altered (Fig. 4). This zooid profiling technique has the advantage over randomly sampling zooids for MART estimation in that recording continuous changes in zooid sizes allows the demonstration of annual growth increments and insights on growth rates and colony longevity (e.g. Fig. 5 of O’Dea & Jackson 2002). Table 1 summarises the set of rules that must be met when conducting either random or systematic measurements of zooids for zooid Fig. 2. Floridina regularis. Scanning electron micrograph of a size MART analysis. These rules reflect the various cri- colony from the Yorktown Formation (Chuckatuck, Virginia, teria mentioned above. USA) showing length (L) and width (W, 150 µm) of a zooid. Measurements are based on maximum dimensions of zooids that meet the criteria for zooid size mean annual range of temperature (MART) analysis, see Table 1 EVIDENCE FOR THE RELATIONSHIP BETWEEN TEMPERATURE AND ZOOID SIZE O’Dea & Jackson (2002) later developed an alterna- tive method for zooid size MART analysis. Instead of The zooid size MART approach is founded upon the conducting random sampling of 20 zooids per colony, negative relationship between zooid size and tempera- they undertook highly systematic sampling of zooids ture that has been demonstrated in both controlled lab- that met the above criteria. This entailed measuring oratory and field situations and over spatial and tempo- sequential generations of disto-laterally budded zooids ral scales. In the following 4 subsections we review and in cupuladriid bryozoans (Fig. 4). They characterised evaluate the growing body of evidence that supports 4 such zooid profiles per colony and found that the this relationship. mean maximum and minimum values correlated well with the MART experienced by the colonies living in contrasting seasonal regimes. This zooid profiling approach requires particular care to avoid measuring the

Fig. 4. Cupuladria aff. biporosa. Zooid size profiling. Scan- Fig. 3. Floridina regularis (as in Fig. 2). Zone of astogenetic ning electron micrograph. White lines = paths of 4 profiles change. Scanning electron micrograph of ancestrula (A) and from the central ancestrula to the colony margin. Dots = region of early colony growth showing a gradual increase in zooids chosen for measurement according to strict rules as zooid size throughout the first few generations. Scale bar = outlined in text. Scale bar = 1 mm. Reprinted from O’Dea 250 µm & Jackson (2002) with permission from Elsevier 138 Mar Ecol Prog Ser 430: 133–146, 2011

Laboratory studies 5° reduction in temperature resulted in a 25% increase in zooid surface area. When normal, warmer tempera- Controlled laboratory studies have consistently tures were restored, new zooids reverted to the smaller demonstrated that zooids increase in size at lower tem- size associated with 29°C. peratures. Menon (1972) showed decreases in both the lengths and widths of zooids of Electra pilosa and Conopeum reticulum as laboratory temperatures Growth in the field increased (temperature regimes: 6, 12, 18 and 22°C for E. pilosa; 12, 18 and 22°C for C. reticulum). Hunter & Temperature-induced variation in zooid size has Hughes (1994) found that Celleporella hyalina pro- been directly investigated in the field in the encrusting duced significantly smaller zooids at 18°C than at 8°C species Conopeum seurati. This was achieved by mea- and that this response occurred irrespective of food suring the maximum lengths and widths of focal supply (10 vs 100 cells ml–1 of Rhodomonas baltica). A colonies that colonised glass slides on 19 occasions later study provided further and independent evidence over 15 mo in the Severn Estuary and conducting for an inverse relationship between temperature and simultaneous measurements of temperature, salinity zooid size in C. hyalina, with autozooids being smaller and food availability, as estimated from chlorophyll a when colonies were reared at 10°C than at 18°C, even (chl a) concentration (O’Dea & Okamura 1999). Sam- when the partial pressure of oxygen was altered from pling intervals were approximately biweekly during normal to hypoxic conditions (Atkinson et al. 2006). periods of rapid growth but less frequent in winter This study also demonstrated that, like length and when growth slowed considerably. General linear width, autozooid volume varies inversely with temper- model analysis revealed that temperature consistently ature. Amui-Vedel et al. (2007) found that Cryptosula accounted for most of the variation in zooid size pallasiana produced longer and wider zooids at 14°C (40.5%) with larger sizes occurring at lower tempera- than at 18°C under equal food concentrations (100 cells tures (e.g. Fig. 1). Salinity, colony identity (genotype) ml–1 of R. baltica). The aforementioned studies were all and an interaction between temperature and salinity conducted on temperate encrusting species that were also influenced zooid size (by 21.1, 3.2 and 22.0%, growing on glass or plexiglass slides whose flat sur- respectively). Factors that had no significant effect on faces will minimise variation in zooid dimensions aris- zooid size were food availability (chl a concentration), ing from topographic complexity. colony growth rate and the reproductive status of col - O’Dea et al. (2007) took advantage of the naturally onies based upon the presence or absence of embryos, high rates of colony cloning in the free-living, tropical eggs or oocytes (O’Dea & Okamura 1999). species Cupuladria exfragminis to observe the effects Other studies that have measured zooid size varia- of temperature upon zooid size amongst genetically tion in the field were conducted on colonies estab- identical clones. Colonies from the Gulf of Panama lished on natural substrata. Lombardi et al. (2006) were halved and the resulting clonal replicates ex- demonstrated that the upright bifoliate species Penta- posed in culture to either 29°C, the normal tempera- pora fascialis produces larger zooids during the colder ture for the Gulf of Panama, or 24°C, a temperature winter than during the warmer summer near Plymouth commonly observed during episodes of upwelling. The (UK) and Tino Island in the Mediterranean. In contrast,

Table 1. Rules for choosing zooids, colonies and species appropriate for zooid size mean annual range of temperature (MART) analysis on cheilostome bryozoans

Zooids Colonies Species

1. Must be autozooids (not kenozooids, vibracula, 1. With >30 ontogenetically 1. Should possess clear auto- avicularia, etc.) complete autozooids, preferably zooid margins 2. From outside zone of astogenetic change many more 2. Autozooid margins are not 2. Not growing on highly obscured by expansion of 3. Not abnormal in size/shape (due to e.g. physical irregular surfaces polymorphs (e.g. avicularia) or damage, biotic interaction, position at or just distal 3. Growth not impeded by frontal budding to bifurcation in budding series or local regeneration competition with other sessile 3. Not distorted by e.g. drying following colony fragmentation) organisms or epibionts 4. Colonies can achieve large 4. From basal series (not frontally budded) 4. Replication of colonies within size (not ‘spot’ colonies with 5. EITHER: Randomly choose 20 autozooids con- species (≥5) determinate growth to small forming to zooid rules 1–3 size) 6. OR: Measure successive generations of autozooids conforming to zooid rules 1–3 (zooid proﬁling) Okamura et al.: Environmental reconstruction by zooid size analysis 139

the average zooid lengths of Cryptosula pallasiana investigated how zooid densities in a colony of the from south Wales were significantly longer in July than upright bifoliate species Pentapora foliacea related to in January, although there was no significant differ- stable oxygen isotope data gathered during a previ- ence in zooid width (Amui-Vedel et al. 2007). These ous investigation by Pätzold et al. (1987). The study field results counter the zooid size–temperature rela- demonstrated covariation between zooid density and tionship that was observed for C. pallasiana in con- contemporaneous δ18O values derived from analyses of trolled laboratory studies (see ‘Laboratory studies’), zooid skeletal walls, such that lower densities, and implying that factors other than temperature may have hence larger zooid sizes, were associated with higher influenced zooid dimensions. O’Dea & Jackson (2002) δ18O values indicative of cooler waters (O’Dea 2005). analysed the growth of free-living cupuladriid bryozoans from tropical American regions that experienced strong seasonal upwelling with closely related Geographical variation in the present day species from non-upwelling environments. They adopted the zooid size profiling approach to charac- Geographical variation in zooid sizes has been noted terise the sizes of zooids in successive generations in a for the encrusting, cave-dwelling species Haplopoma colony (Fig. 4). Strong cyclical patterns of increasing sciaphilum, which produces large zooids in Sweden zooid sizes were characteristic of colonies that experi- where it is colder, smaller zooids in the North Adriatic enced the seasonal flux of cold water associated with where it is warmer, and yet smaller zooids in France upwelling, when temperatures can drop by >10°C where it is warmest (Silén & Harmelin 1976). Similarly, within 1 wk (D’Croz & O’Dea 2007; see also Fig. 1). In mean zooid lengths of Pentapora fascialis showed a contrast, colonies from non-upwelling environments significant and negative correlation with mean annual revealed no significant variation in zooid size through- temperature for colonies collected from 9 sites in out their zooid size profiles. the Mediterranean and the UK (Lombardi et al. 2006). In addition to those studies that compared zooid size Conversely, Novosel et al. (2004) found no systematic directly with temperature, there are several that have differences in zooid lengths between 2 sites in the done so indirectly, including an early observation that North and South Adriatic that could be explained by led to the development of the zooid size MART temperature. Their study was highly inclusive, ana- approach (Okamura 1987). Autozooids of Electra pilosa lysing 14 cheilostome species. However, the sites were were observed to vary in size over the year in the each characterised by varying and complex environ- Menai Straits, North Wales. The evidence was derived mental regimes, including fluctuating salinities at 1 by dividing the total number of zooids per colony by site due to large freshwater inputs via the Zrmanja colony size, allowing an estimate of mean zooid size River and an underground system of karst canals per colony at different times of the year (Okamura acting as temporary freshwater springs. Substantial 1987). Zooid size appeared to be inversely related to fluctuations in salinity are likely to have confounded temperature, with smaller zooids produced during the the zooid size response to temperature (see ‘Confound- warmer summer months. There was no apparent link ing factors’ below). Furthermore, the wholly inclusive between zooid size and productivity (e.g. spring and random selection of zooids for measurement, without autumn phytoplankton blooms in the Menai Straits; due care, may have resulted in measures of zooids with Jones & Spencer 1970, Al-Hasan et al. 1975). aberrant growth or those from the zone of astogenetic Other studies have taken a related approach using change. This may explain why the authors were zooid densities as a proxy for size. O’Dea & Okamura required to adopt a non-parametric approach to analy- (2000a) exploited the production of annual growth ses since occasional large variances in zooid length lines in the upright bifoliate species Flustra foliacea to would be expected if early stage or distorted zooids temporally constrain measures of zooid densities in were inadvertently measured. colonies from Wales, Denmark and Nova Scotia. They found that zooid densities varied cyclically throughout the year in apparent synchrony with seasonal variation Deep-time variation in temperature, such that the lowest densities and hence the largest zooids occurred during the cooler Environments vary not only over space but also over spring and autumn periods. Records of planktonic time. The first study to explicitly use zooid size in bryo - primary productivity, in particular the bimodal spring zoans to estimate relative changes in seawater temper- and autumn phytoplankton blooms that characterise atures over geological time scales focused on 8 species the Menai Straits and the Skagerrak (Pettersson 1991, common to both the Pliocene Coralline Crag Forma- Blight et al. 1995), indicated that food availability could tion in the eastern UK and in the present-day waters not have significantly affected zooid size. O’Dea (2005) around the UK (Okamura & Bishop 1988). Specimens 140 Mar Ecol Prog Ser 430: 133–146, 2011

in the collections of the Natural History Museum, Lon- mura 1987, O’Dea & Okamura 1999) nor with variation don, were identified that provided at least 5 autozooids in zooid density in E. pilosa, C. reticulum or Celle- for measurement that were unobstructed in growth porella hyalina in laboratory experiments (Menon and outside the zone of astogenetic change. Mean 1972, Hunter & Hughes 1994) or in Flustra foliacea in zooid sizes were significantly smaller in 5 of the 8 spe- the field (O’Dea & Okamura 2000a). However, analysis cies in the Coralline Crag than the present day, corro - of morphological responses of E. pilosa in controlled borating previous estimates (see Okamura & Bishop laboratory studies demonstrated that food concentra- 1988 for references) that the Coralline Crag was de - tion produced predictable yet non-linear effects on posited in a sub-tropical sea considerably warmer than zooid size (Hageman et al. 2009). At very low food today (see also Williams et al. 2009 for further evalua- levels, stunted colonies with small zooids developed. tion of the Coralline Crag environment). Replicating At low to intermediate food concentrations, zooid sizes the same methods and many shared species, A. O’Dea increased with food levels up to a threshold food con- (unpublished data), found that zooid sizes in the centration (~7.51 µg l–1 chl a) where maximum zooid younger Pleistocene Red Crag formation were midway sizes were produced. At food concentrations above this between those of the Coralline Crag and modern day, threshold, zooid sizes diminished and then stabilised at corroborating the inferred slightly warmer conditions a size smaller than the maximum size observed at the of the Red Crag relative to the present day and cooler threshold. The magnitude of food-induced variation in conditions relative to the Coralline Crag (Head 1998). zooid size was associated with these zooid size trends. In contrast, Berning et al. (2005) found that zooid areas Thus, maximum variance was coincident with the were notably smaller in the Late Miocene cheilostomes maximum size, and minimum variance was observed from the putatively cooler Guadalquivir Basin in the for zooids of submaximum size above the threshold eastern Atlantic than they were in the same species food concentration. Intermediate levels of variation in 2 warmer and nearly coeval Mediterranean assem- were associated with zooids experiencing low to blages, suggesting that factors other than temperature threshold food concentrations. may have influenced size (e.g. freshwater input from If the responses of Electra pilosa apply broadly, they the Guadalquivir River). imply that food-induced variation in zooid size will be minimal above some threshold food concentration while greater variation in zooid size in response to food Confounding factors levels can be expected at the threshold food concentration and at levels below. These latter concentrations When applying the zooid size MART technique, it are estimated to be within the range of concentrations should always be borne in mind that water tempera- reported for the natural environment and typical of ture can be influenced by factors other than seasonal waters from which the E. pilosa material was originally variation, such as upwelling or depth. In these cases, collected (0.256 to 16.0 µg l–1 chl a; Hageman et al. while the bryozoan responses should represent a true 2009). This suggests that variation in food levels may at indication of temperatures experienced, the minimum times confound temperature effects by enhancing or and maximum zooid sizes may reflect temperature reducing seasonal temperature effects. Previous stud- variation as a result of ocean currents. Systematic sam- ies that have reported no effect of food either used food pling of species that produce annual growth lines may concentrations that were higher than the threshold provide a means of recognising multiple drivers of concentration for E. pilosa, or they did not report food zooid size. For instance, this may be the case if succes- levels. Hageman et al. (2009) therefore speculated that sive zooid generations demonstrate >1 peak in size studies revealing no food effect may have been con- over the course of a single year. ducted well above the critical value to invoke variation Intracolonial zooid size can also be influenced by in zooid size. other environmental factors besides temperature, e.g. The study by Hageman et al. (2009) has implications by distortion arising from microenvironmental varia- for inferences of temperature regimes made by the tion such as irregularities of the substratum over which zooid size MART approach, with variation in food colonies are growing (Boardman et al. 1969) or follow- levels potentially confounding inferred regimes. How- ing injury or abrasion. More pervasive environmental ever, there are several caveats. (1) It is not clear how influences on zooid size include variation in food avail- food-induced changes in zooid size in Electra pilosa ability, salinity or flow regimes. when fed an algal monoculture might equate to food- There are conflicting results regarding the influence induced changes in zooid size when a diversity of dif- of food on zooid dimensions. For instance, food avail- ferent food types is available in natural environments. ability was not associated with variation in zooid size in (2) Before the experiment is repeated on other species, Electra pilosa and Conopeum seurati in the field (Oka- it is impossible to discount the possibility that the trends Okamura et al.: Environmental reconstruction by zooid size analysis 141

observed for E. pilosa may not be universal. (3) Changes boundary layer, thereby avoiding excessive flows that in food concentrations from 103 to 104 cells ml–1 (esti- are detrimental to feeding (Okamura 1985, Eckman & mated as equivalent to 1.251 to 12.51 µg l–1 chl a) and Okamura 1998). from 104 to 105 cells ml–1 (12.51 to 125.1 µg l–1 chl a) Finally, a number of studies have demonstrated that were associated with +14.8% and –2.7% changes in zooid size variation is influenced by genotype. This zooid area, respectively. These effects are much smaller has been shown in laboratory studies of Celleporella than the effect of temperature on Conopeum seurati, hyalina and Electra pilosa (Hunter & Hughes 1994, which explained 40.51% of the variation in zooid size Atkinson et al. 2006, Hageman et al. 2009). Genotype when accounting for variation in food levels ranging has also been inferred to influence zooid size amongst from near 0 to ~45 µg l–1 chl a (O’Dea & Okamura sexually produced colonies that developed under field 1999). conditions, accounting for 3.16% of the variation in As discussed earlier, oxygen may also influence zooid size of Conopeum seurati (O’Dea & Okamura zooid size and indeed may provide a mechanistic 1999). Such variation dem onstrates the variation in explanation for the temperature–size rule. The major- thermal reaction norms amongst individuals referred ity of shallow water and shelf environments that sup- to earlier, and which, for instance, would allow selec- port bryozoan growth can generally be expected to be tion and conformation to Bergmann’s Rule. near oxygen saturation at any given temperature due to atmospheric mixing and photosynthesis. This indi- cates that the systematic variation between seawater ZOOID SIZE MART APPROACH: APPLICATIONS temperature and oxygen content should not be con- AND INSIGHTS founded by e.g. oxygen depletion as a result of biological processes that might indirectly influence zooid The zooid size MART approach has been applied to size. However, the effects of salinity are potentially several situations to interpret palaeoenvironmental con- more complicated due to the inverse relationship be - ditions. Estimates of MART from encrusting cheilo - tween oxygen concentration and salinity levels. This stomes from the Pliocene Coralline Crag and the relationship prompted O’Dea & Okamura (1999) to Miocene ‘Faluns’ (O’Dea & Okamura 2000b) suggest conclude that salinity, probably as a result of its effect that northwest European seas were considerably more on oxygen solubility, significantly influenced zooid size equable (less seasonal) than they are today, a conclu- in Conopeum seurati, accounting for 21.13% of the sion upheld by open ocean palaeoenvironmental prox- variation in zooid size. However, Spicer & Gaston ies and oceanic modelling from these times (e.g. (1999) highlighted that oxygen partial pressure drives Cronin & Dowsett 1996, Lécuyer et al. 1996, Knowles the diffusion gradient in organisms (along with respi- et al. 2009 and references therein). More recently, the ratory pigments). At higher salinities, oxygen solubility zooid size MART approach has been applied for the will be reduced, but the oxygen partial pressure first time across broad spatial scales focusing on the should not be affected. Thus, the changes in zooid size Pliocene North Atlantic, especially on the time in and in C. seurati (21.13%) may be caused by the direct around the mid-Pliocene warm period (Knowles et al. effect of salinity rather than the covariance of salinity 2009). Cheilostome assemblages from the Coralline and oxygen solubility (or concentration). In any case, Crag Formation (UK), the Yorktown Formation (Vir- the fact that O’Dea & Okamura (1999) retrieved tem- ginia, USA), the Duplin Formation (South Carolina, perature as the most significant factor in driving zooid USA), the Lower Tamiami Formation (Florida, USA) size even under temperate estuarine conditions sug- and the Cayo Agua Formation (Panama) were used to gests that salinity effects may rarely swamp tem - reconstruct MART estimates and to investigate pat- perature effects if temperature and salinity generally terns of heat transferral from equatorial to mid-latitude covary. regions. Knowles et al. (2009) corroborated previous Flow regime can also influence zooid size. A field evidence for upwelling in the southern Caribbean and transplant study in the relatively simple hydrodynamic in Florida. Their study also indicated that the warm regime of the Rapids at Lough Hyne, Ireland, demon- current flowing northeast from the Caribbean resulted strated that zooids of Membranipora membranacea in reduced seasonality along the eastern seaboard of decrease in size with increasing flow (Okamura & Par- the USA, and that it was deflected across the Atlantic tridge 1999). The equivalent growth rates of colonies further north from the Cape Hatteras area of North regardless of flow provided evidence that flow- Carolina where it is deflected today. The study also induced miniaturisation may enable effective suspen- showed that seasonal variation in the southern North sion feeding by creating favourable flow microenvi- Sea was greater than that experienced today. ronments. This may be achieved by size alterations Free-living cupuladriid bryozoans have a rich fossil that place lophophores into slower flow regimes of the record in the Caribbean extending back to the time 142 Mar Ecol Prog Ser 430: 133–146, 2011

before the Isthmus of Panama closed and have been Another study used a modified version of the zooid used as key proxies for inferring environmental size MART approach to reconstruct MART from fossil change during the closure of the Isthmus via the zooid cheilostome assemblages of 2 consecutive units of the size MART approach. Today, seasonal upwelling lower Setana Formation (1.2 to 1.0 Ma, Kuromatsunai, brings cool, nutrient-rich waters along the Pacific coast Hokkaido, Japan; Dick et al. 2007). A ‘specimen- for 3 mo each year (D’Croz & O’Dea 2007). In contrast, limited MART’ (SL-MART) approach was developed no upwelling occurs along the Caribbean coast (D’Croz that simulates additional MART estimates from avail- & Robertson 1997). The zooid size MART approach able data to enable use of small datasets when few was initially applied to cupuladriids from the present- specimens are available for study. The results sug- day by profiling the sizes of zooids from the centre of gested more pronounced seasonality in the lower than colonies to their margins (Fig. 4; O’Dea & Jackson in the upper Setana Formation, but it was concluded 2002). Strongly fluctuating patterns in zooid size were that greater sample sizes were advisable for recon- documented in colonies from the upwelling Gulf of struction of palaeoseasonality. The SL-MART tech- Panama but not in geminate species from the non- nique is therefore perhaps best used as a means of upwelling Caribbean coast, and resulting estimates of gaining preliminary insights into palaeoenvironmental MART obtained were within an accuracy of ±1°C of conditions. actual MARTs as derived from multiple oceanographic In a very recent study, Knowles et al. (2010) undertook data (see O’Dea & Jackson 2002 for references). Subse- the first investigation to examine how results from the quent estimates of MART from nearly 150 fossil cupu- zooid size MART approach and stable isotope analyses ladriid colonies suggest that strong seasonal upwelling relate to the actual measured ranges of temperature ex- was a permanent feature of the Caribbean when the perienced by cheilostome bryozoans as recorded by a inter-oceanic Central American Seaway connected the datalogger. They found that the MART implied by zooid Pacific and Caribbean (O’Dea et al. 2007). The end of size variability in Pentapora foliacea (overall mean of upwelling in the Caribbean occurred rapidly during 6.8 and 6.9°C at 2 sites in Wales) gave a good approxima- the final stages of isthmus closure, even though the for- tion to the recorded range in temperature (overall mean mation of the isthmus was a slow geological process range = 7.8°C, based on a mean minimum recorded (Coates & Obando 1996). When the Caribbean became temperature of 8.2°C and a mean maximum recorded isolated from the Tropical Eastern Pacific, there was a temperature of 16.0°C), while the temperature ranges synchronous shift from heterotrophic-dominated to reconstructed by oxygen stable isotopes were narrower. auto- and mixotrophic-dominated benthic communi- The latter result appeared to reflect secondary skeletal ties, consistent with a rapid decline in planktonic pro- thickening that homogenised the temperature signal by ductivity and nutrient levels. The high-resolution esti- time-averaging. How ever, the good approximation to mates of upwelling derived from the zooid size MART maximum recorded temperatures by oxygen stable approach have revealed that ‘nutriphilic’ coral, bry- isotopes (range = 14.8 to 16.9°C) demonstrated that zooid ozoan and molluscan taxa went extinct some 1 to 2 Ma size variation and stable isotope analyses can provide after the end of upwelling, thus challenging the con- independent and valuable proxies for inferring tempera- ventional wisdom that cause and effect coincide in ture regimes,with data from stable oxygen isotopes relat- deep-time. ing MART values derived from zooid size variation to Polar material has been the focus of recent zooid size absolute temperatures. MART analysis. Clark et al. (2010) undertook a pilot study to examine cheilostomes from the Early Plio- cene Weddell Sea, Antarctica. Although relatively ADVANTAGES AND LIMITATIONS OF THE ZOOID few colonies were found to meet criteria for SIZE MART APPROACH analysis, the material provided MART estimates (range = 4.8 to 10.3°C) that were consistently greater The zooid size MART approach offers several advan- than the seasonal variation in temperature in the pre- tages. (1) It represents a relatively inexpensive and sent day in the Weddell Sea (2°C; Whitehouse et al. quick method to infer palaeoclimate regimes. (2) MART 1996). Although zooid size MART estimation does not values can be used directly as estimates of temperature provide an indication of absolute temperatures, the variation in environments from which the individual results from this investigation suggest that summer colonies were collected. In contrast, estimates of mean maximum temperatures may have approached or zooid size provide no environmental information on exceeded 8°C in the Early Pliocene (given a minimum their own but require comparisons with mean zooid of –1.8°C when seawater freezes), similar to the pre- sizes of colonies from different environments (e.g. as in sent-day seasonal temperature variation in coastal sur- Okamura & Bishop 1988, Berning et al. 2005, Lombardi face waters of southern Patagonia. et al. 2006). (3) The approach can be used to charac- Okamura et al.: Environmental reconstruction by zooid size analysis 143

terise the ecology of present-day en vironments that RECOMMENDATIONS FOR FUTURE STUDIES are difficult to access or for which relevant temperature data are unavailable. (4) The zooid size MART The foregoing discussion has reviewed the develop- approach provides information on the environment ment and application of the zooid size MART approach experienced by an individual colony that can be and the negative relationship between zooid size and related to the life history of that individual colony (e.g. temperature that provides the foundation for the ap- allocation to reproduction, defense and growth), proach. Here we suggest future areas for research to thereby enabling direct correlation of life history address outstanding questions and to identify exten- responses with environmental regimes. We reiterate sions and further developments of the zooid size MART that bryozoans are the only colonial invertebrates that approach. enable such insights as a result of their unique combi- Further exploration of the mechanisms that underlie nation of traits: intracolonial polymorphism, the pro- the zooid size–temperature relationship and its gener- duction of carbonate skeletons and determinate zooid ality is merited. Atkinson et al. (2006) have clearly sizes. demonstrated that zooid sizes in Celleporella hyalina Like any technique, the zooid size MART approach can be influenced by both temperature and oxygen has limitations that are important to appreciate. (1) It levels. Are such responses general amongst cheilo- requires appropriate material for analysis—a necessity stome bryozoans? Is there an adaptive basis for the size inherent to all methods of assessing temperature change, and how might understanding this help us to regimes. (2) Adherence to the set of rules outlined interpret size variation over time and space? Their in Table 1 is critical, since violating assumptions of finding that colony volumes do not conform to the tem- the approach is likely to give misleading results. For perature–size rule is intriguing, as it suggests that instance, ignoring the strict criteria for measurement, zooids, but not colonies, are the units that show equiv- such as measuring distorted zooids or zooids within the alent responses to those displayed by solitary organ- zone of astogenetic change, will compromise infer- isms. Does this vary amongst species according to their ences. We have extensively explored how factors other degree of colony integration or mode of life? Do zooids than temperature may influence zooid size, concluding in poorly integrated colonies (e.g. uniserial runners that these are both less pervasive and exert weaker that lack zooid polymorphism) show stronger tempera- effects than temperature. (3) It assumes that colonies ture-induced variation in zooid size than zooids in grow throughout the year (or that growth spans the highly integrated colonies (multiserial forms with a period of maximum and minimum temperatures). Lack high degree of zooid polymorphism)? of growth during, for instance, periods of low food The body of evidence to date suggests that chei - availability (e.g. in winter) may result in an inferred lostome zooids generally respond to temperature. MART that is lower than the actual temperature. Never theless, phylogeny can influence many or- There are various means of recognising when in- ganismal responses (Harvey & Pagel 1991), and ferences for MART may be compromised, including temperature-induced zooid size variation has been aberrant estimates obtained from a small number of demonstrated for relatively few cheilostome species. colonies. For instance, a species that consistently estimates a smaller MART relative to other species may be ceasing growth during winter periods. This could be indicated by the presence of annual growth check lines (Fig. 5), and such material could then be avoided. Also, zooid size changes resulting from variation in salinity could be misinterpreted to represent temperature effects on size. In environments where temperature and salinity covary, such as in temperate estuarine conditions, temperature appears to exert an overwhelming effect on zooid size variation (O’Dea & Okamura 1999). However, salinity could be confounding in environments where temperatures remain relatively constant. For instance, coastal salinities may vary due to seasonal rainfall in the tropics or ice melting in polar regions. In such cases, assemblage information may help to inform Fig. 5. Melicerita sp. Growth cessation during the winter. Scanning electron micrograph of a fragment from the Coral - on the type of environment in which bryozoans lived, line Crag Formation (Suffolk, UK) showing skeletal thicken- with MART values unexpected for such environments ing typical of an annual growth check line (white arrows). suggestive of possible salinity effects. Scale bar = 250 µm 144 Mar Ecol Prog Ser 430: 133–146, 2011

Although the apparent ubiquity of the temperature– CONCLUSION size rule suggests that phylogeny will play a minor role, further investigation is warrented. If phylogenetic Our review demonstrates a strong and growing history proved important in determining how zooids body of evidence that the zooid size MART approach respond to temperature, then more strategic sampling is a unique and independent proxy for environmental and analyses would be advisable. For instance, ana- reconstruction. The above-described investigations of lyses could be adopted to deal with correlated evolu- bryozoan growth in laboratory and field conditions tion as a result of common phylogenies (Harvey & are nearly unanimous in confirming the inverse rela- Pagel 1991), and taxa could be targeted or avoided to tionship between zooid size and temperature. Fur- minimise phylogenetic constraints (Gould & Lewontin thermore, the apparent ubiquity of the temperature– 1979). size rule lends additional support. Temperature con- Exploring whether an approach based on tempera- sistently provides, either directly or indirectly, a per- ture-induced variation in module size can be extended vading and dominant influence on zooid size. The to other groups could prove to be highly productive potential for other environmental factors to influence and might provide key insights on very ancient envi- size should nevertheless always be borne in mind ronments. Cyclostome bryozoans are the most obvious when using the zooid size MART approach. How- candidate for such exploration, although their mode ever, incorporating other proxies may help to deal of growth would require focusing on different charac- with such confounding factors, and palaeoenviron- ter(s) for measurement (e.g. the aperture). If cyclo - mental reconstruction is considerably strengthened stomes proved applicable, the use of the zooid size when multiple independent proxies are used. Chei- MART approach to reconstruct ancient environments lostomes may provide further potential in this might be extended from the Upper Jurassic (155 Ma; respect, since studies indicate the feasibility of com- when cheilostomes originated) to the Lower Ordovician bining the zooid size MART approach with stable iso- (480 Ma; when cyclostomes originated). tope analyses of the same specimens (O’Dea 2005, Finally, refinement of the zooid size MART approach Knowles et al. 2010), provided the mineralogy of the might enable more accurate and powerful estimates of specimens has been well constrained (Smith et al. temperature regimes (Dick et al. 2007). This could cer- 2004) and diagenesis is absent (T. Knowles et al. tainly be achieved by increasing the number of data unpubl. data). points in the model to incorporate more species and to We conclude that the zooid size MART approach extend geographic cover. It could be argued that more represents a robust proxy for environmental recon- accurate estimates might be gained by more precise struction that warrants equal consideration for use as measurements of zooid size. For instance, a pilot study that given to traditional proxies such as stable isotope confirmed that greater precision can be gained through analyses or alkenones. In addition, because the zooid length and width measurements obtained from scan- size MART approach entails sampling over discrete ning electron microscopy (SEM) since SEM images periods of time, insights into the actual annual tem- can reduce error in size estimation relative to mea - perature range are possible. In this respect, the surements made using a stereomicroscope (Knowles approach offers equal or greater precision than analy- 2009). However, a requirement to use SEM images for ses of traditional proxies based on combined samples measurement is not warranted for 3 reasons. (1) It deriving from a number of years (e.g. geochemical would then be advisable to develop a new model using signals from foraminifera) and which may be further SEM-based measurements. (2) A highly attractive compromised by bioturbation. Furthermore, because aspect of the zooid size MART ap proach is that it is it is a relatively inexpensive and quick method, the easy, inexpensive and requires only standard labora- zooid size MART approach can provide a means of tory equipment. (3) The reliability (e.g. regression gauging environmental variation and of identifying model with 95% confidence limits within ±1°C) and where or when more expensive proxies might be accuracy (e.g. concordance with known temperature adopted. It therefore merits broad recognition by regimes or other proxies) of the zooid size MART environmental and evolutionary biologists and cli- approach appears to us very reasonable in view of mate modellers. Finally, its incorporation in multi- background noise inherent to both new data and the proxy toolkits used in palaeoenvironmental research model. We note, however, that SEM images can be will enable more robust interpretations. Ultimately, very helpful to delineate zooid boundaries, particularly such multiproxy-based research with strategic focus in fossil specimens (e.g. Knowles et al. 2009), and for on palaeoclimate change will enable better under- this reason, making measurements on SEM images standing and prediction of how the earth system has can be highly justified as a means of improving data responded in the past and how it may therefore collection. respond to future climate change. Okamura et al.: Environmental reconstruction by zooid size analysis 145

Acknowledgements. We thank P. Taylor for support in Isthmus. In: Jackson JBC, Budd AF, Coates AG (eds) the development and application of the zooid size MART Evolution and environment in tropical America. University approach and for greatly facilitating our research over the of Chicago Press, Chicago, p 57–75 years; M. Williams, a non-bryozoologist who recognised the D’Croz LD, O’Dea A (2007) Variation in upwelling along the potential of the zooid size MART approach and has enthusias- Pacific shelf of Panama and implications for the distribu- tically championed its use; R. Hughes and J. Jackson for inspi- tion of nutrients and chlorophyll. Estuar Coast Shelf Sci ration and insights on the evolutionary ecology of modular 73:325–340 animals; S. Hageman for discussion, R. Dewel for the SEM D’Croz LD, Robertson DR (1997) Coastal oceanographic con- image used for Fig. 1; and 3 reviewers for comments that ditions affecting coral reefs on both sides of the Isthmus of helped to improve our manuscript. Our research has obtained Panama. Proc 8th Int Coral Reef Symp 2:2053–2058 funding and support from the Natural Environment Research Dick MH, Hirose M, Takashima R, Ishimura T, Nishi H, Council, the Biotechnology and Biological Research Council, Mawatari SF (2007) Application of MART analysis to infer the Smithsonian Tropical Research Institute and the Natural paleoseasonality in a Pleistocene shallow marine benthic History Museum. environment. In: Okada H, Mawatari SF, Suzuki N, Gau- tam P (eds) Proc Int Symp ‘Origin and Evolution of Natural Diversity’, 1–5 October 2007, Sapporo, p 93–99 LITERATURE CITED Eckman JE, Okamura B (1998) A model of particle capture by bryozoans in turbulent flow: significance of colony form. 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Submitted: April 27, 2010; Accepted: November 30, 2010 Proofs received from author(s): March 1, 2011